OBJECTIVE

We sought to determine whether 4000 IU/d (vs 2000 IU/d) of vitamin D during pregnancy is safe and improves maternal/neonatal 25-hydroxyvitamin D [25(OH)D] in a dose-dependent manner.

METHODS

A total of 257 pregnant women 12-16 weeks' gestation were enrolled. Randomization to 2000 vs 4000 IU/d followed 1-month run-in at 2000 IU/d. Participants were monitored for hypercalciuria, hypercalcemia, and 25(OH)D status.

RESULTS

Maternal 25(OH)D (n = 161) increased from 22.7 ng/mL (SD 9.7) at baseline to 36.2 ng/mL (SD 15) and 37.9 ng/mL (SD 13.5) in the 2000 and 4000 IU groups, respectively. While maternal 25(OH)D change from baseline did not differ between groups, 25(OH)D monthly increase differed between groups (P < .01). No supplementation-related adverse events occurred. Mean cord blood 25(OH)D was 22.1 ± 10.3 ng/mL in 2000 IU and 27.0 ± 13.3 ng/mL in 4000 IU groups (P = .024). After controlling for race and study site, preterm birth and labor were inversely associated with predelivery and mean 25(OH)D, but not baseline 25(OH)D.

CONCLUSIONS

Maternal supplementation with vitamin D 2000 and 4000 IU/d during pregnancy improved maternal/neonatal vitamin D status. Evidence of risk reduction in infection, preterm labor, and preterm birth was suggestive, requiring additional studies powered for these endpoints.

Exercise and muscle strength are important determinants of bone mass. Studies were carried out in normal young adult white males to determine the effects of exercise on vitamin D and mineral metabolism. Fourteen men who had engaged in regular muscle-building exercises for at least 1 year and 14 age-matched controls (age range, 19-36 year) were hospitalized on a metabolic ward and were given a constant daily diet estimated to contain 400 mg of calcium, 900 mg of phosphorus, 110 mEq of sodium, 65 mEq of potassium, and 18 mEq of magnesium. Body weight averaged 78 +/- 3 kg in the exercisers and 72 +/- 2 kg in the controls (NS). Serum calcium, ionized calcium, phosphate, magnesium, somatomedin-C, and immunoreactive parathyroid hormone (PTH) were not different in the two groups, whereas serum Gla-protein (39 +/- 5 vs. 24 +/- 2 ng/ml, p less than 0.01), 25-hydroxyvitamin D (23 +/- 2 vs. 16 +/- 2, p less than 0.05) and 1,25-dihydroxyvitamin D [1,25(OH)2D] (40 +/- 2 vs. 29 +/- 2 pg/ml, p less than 0.01) were higher in the exercisers than in the controls. Urinary calcium, phosphorus, sodium, potassium, creatinine clearance, and norepinephrine were not different in the two groups, whereas urinary magnesium (12.6 +/- 1.0 vs. 9.4 +/- 0.5 mEq/d, p less than 0.01) and urinary cyclic adenosine 3',5'-monophosphate (cyclic AMP) (2.52 +/- 0.19 vs. 1.72 +/- 0.20 nM/dl glomerular filtrate, p less than 0.01) were higher in the exercisers than in the controls.(ABSTRACT TRUNCATED AT 250 WORDS)

The influence of a high-protein meal as compared to fasting on the disposition of simultaneous intravenous and oral doses of propranolol, as well as on indocyanine green clearance, was examined in six normal subjects. The intravenous dose (0.1 mg/kg) was unlabeled propranolol and the oral dose (80 mg) was a stereospecifically deuterium-labeled pseudoracemate of propranolol. Systemic clearance of propranolol increased 38%, from 1005 +/- 57 to 1384 +/- 115 ml/min (mean +/- SE; P less than 0.05) as a result of the meal, with no change in t1/2 or apparent volume of distribution. A 12% decrease in oral clearance occurred with the meal but was not statistically significant (3717 +/- 185 ml/min, fasting; 3245 +/- 498 after meal), whereas bioavailability increased 67% (27.2% +/- 1.7% fasting; 45.5% +/- 4.3% after meal; P less than 0.01). Estimated hepatic blood flow, as measured by indocyanine green clearance, rose 34% 60 minutes after the meal (1719 +/- 155 ml/min fasting; 2304 +/- 218 ml/min after meal; P less than 0.02). A difference was observed in the oral clearance of the propranolol enantiomers in the fasting state, but this difference was unaffected by the meal. These alterations in propranolol disposition, as the result of a high-protein meal, are consistent with a transient increase in hepatic blood flow.

The influence of a meal on the disposition and metabolism of oral propranolol was examined in six normal subjects. The meal induced a mean 53% increase in propranolol bioavailability (range 2% to 92%; P less than 0.01) without affecting time to maximum concentration, half-life, or the amount of unchanges drug in urine. There was no effect on the plasma concentrations of 4-hydroxypropranolol or four other metabolites. The increased bioavailability was linearly related to the protein content of the meal (r = 0.884, P less than 0.02) above a threshold content of about 7 gm.

OBJECTIVE

Childhood trauma has been associated adult stress-related disorders. However, little is known about physiologic alterations in adults with a history of early life trauma that do not have current psychiatric or medical diagnoses. In this study, the relationships between childhood adversity and cytokine and C-reactive protein (CRP) levels in healthy adults were examined.

METHODS

Participants included men (n = 18) and women (n = 20) who did not meet DSM-IV criteria for Axis I psychiatric disorders or any major medical illness. Cytokine and CRP levels were obtained from baseline blood samples. Subjects completed the Early Trauma Inventory Self Report (ETI-SR). The primary outcomes included serum interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), interleukin-1β (IL1-β), and CRP levels. In addition, the mean numbers of traumatic experiences (sexual, physical, emotional, general, and the summed total) were measured.

RESULTS

Significant positive associations were found between the total ETI score and IL-6 (p = 0.05), IL1-β (p < 0.05), and TNF-α (p = 0.01). Significant positive correlations were found between the number of general traumas and IL1-β (p < 0.05), TNF-α (p < 0.05), and IL-6 (p < 0.01). Neither the total number of traumas nor any of the trauma subscales were significantly associated with CRP levels.

CONCLUSIONS

The positive association between childhood trauma and basal cytokine levels supports the extant literature demonstrating the long-term impact of childhood trauma and stress on homeostatic systems. Importantly, this association was found in healthy adults, suggesting that these alterations may precede the development of significant stress-related psychiatric disorder or disease.

OBJECTIVE

The aim of this pilot study was to generate an initial estimate of the prevalence and correlates of diabetic retinopathy in a racially and ethnically diverse sample of youth with Type 1 and Type 2 diabetes mellitus.

METHODS

A pilot study was conducted among 222 individuals with Type 1 diabetes (79% non-Hispanic white, 21% other) and 43 with Type 2 diabetes (28% non-Hispanic white, 72% other), all of>> 5 years duration (mean duration 6.8 years) who participated in the SEARCH for Diabetes in Youth study. Diabetic retinopathy was assessed using non-mydriatic retinal photography of both eyes.

RESULTS

The prevalence of diabetic retinopathy was 17% for Type 1 diabetes and 42% for Type 2 diabetes (odds ratio 1.50, 95% CI 0.58-3.88; P = 0.40 adjusted for age, duration, gender, race/ethnicity, parental education and HbA(1c). HbA(1c) was significantly higher among those with any diabetic retinopathy (adjusted mean 79 mmol/mol, 9.4%) vs. no diabetic retinopathy (adjusted mean 70 mmol/mol, 8.6%) (P = 0.015). LDL cholesterol was also significantly higher among those with any diabetic retinopathy (adjusted mean 107.2 mg/dl) compared with those without diabetic retinopathy (adjusted mean 97.9 mg/dl) (P = 0.04).

CONCLUSIONS

The prevalence of diabetic retinopathy in contemporary young individuals was substantial, particularly among minority youth and those with Type 2 diabetes. Further long-term study of diabetic retinopathy in youth is needed.

Enantioselective hydrolysis of oral racemic methylphenidate (dl-MPH) by carboxylesterase 1 (CES1) limits the absolute bioavailability of the pharmacologically active d-MPH isomer to approximately 30% and that of the inactive l-MPH to only 1-2%. Coadministration of dl-MPH with ethanol results in elevated d-MPH plasma concentrations accompanied by CES1-mediated enantioselective transesterification of l-MPH to l-ethylphenidate (EPH). The present study tested the hypothesis that administration of the pure isomer dexmethylphenidate (d-MPH) will overcome the influence of ethanol on d-MPH absorption by eliminating competitive CES1-mediated presystemic metabolism of l-MPH to l-EPH. Twenty-four healthy volunteers received dl-MPH (0.3 mg/kg) or d-MPH (0.15 mg/kg), with or without ethanol (0.6 g/kg). During the absorption phase of dl-MPH, concomitant ethanol significantly elevated d-MPH plasma concentrations (44-99%; P < 0.005). Furthermore, immediately following the ethanol drink the subjective effects of "high," "good," "like," "stimulated," and overall "effect" were significantly potentiated (P ≤ 0.01). Plasma l-EPH concentrations exceeded those of l-MPH. Ethanol combined with pure d-MPH did not elevate plasma d-MPH concentrations during the absorption phase, and the ethanol-induced potentiation of subjective effects was delayed relative to dl-MPH-ethanol. These findings are consistent with l-MPH competitively inhibiting presystemic CES1 metabolism of d-MPH. Ethanol increased the d-MPH area under the curve (AUC)(0-inf) by 21% following dl-MPH (P < 0.001) and 14% for d-MPH (P = 0.001). In men receiving d-MPH-ethanol, the d-MPH absorption partial AUC(0.5-2 hours) was 2.1 times greater and the time to maximum concentration (T(max)) occurred 1.1 hours earlier than in women, consistent with an increased rate of d-MPH absorption reducing hepatic extraction. More rapid absorption of d-MPH carries implications for increased abuse liability.

1,25-Dihydroxyvitamin D [1,25-(OH)2D] is the principal mediator of the biologic effects of vitamin D. We showed previously that obese white subjects have low serum vitamin D and 25-hydroxyvitamin D (25-OHD) with increased serum-immunoreactive parathyroid hormone (PTH) and 1,25-(OH)2D, low urinary calcium, and increased urinary cyclic adenosine 3',5'-monophosphate (cyclic AMP) compared with nonobese white individuals. To determine whether 25-OHD modulates calcium metabolism, the effects of 25-OHD3, 40-100 micrograms/day for 9 days, were compared in seven obese and seven nonobese white subjects who were between the ages of 20 and 34 years. Each of them was hospitalized on a metabolic ward and given a constant daily diet that contained 400 mg calcium, 900 mg phosphate, and 18 mEq magnesium. Whereas 25-OHD3 increased mean serum 25-OHD from 7 +/- 1 to 37 +/- 5 ng/ml (P less than 0.01) and urinary calcium from 102 +/- 18 to 146 +/- 17 mg/day (P less than 0.001) and decreased mean serum 1,25-(OH)2D from 40 +/- 2 to 28 +/- 2 pg/ml (P less than 0.01) and urinary cyclic AMP from 3.23 +/- 0.57 to 2.00 +/- 0.17 nM/dl GF (P less than 0.05), it did not change mean serum calcium, ionized calcium, phosphate, magnesium, immunoreactive PTH or urinary phosphate, or creatinine clearance in the obese subjects. In contrast, 25-OHD3 increased mean serum 25-OHD from 16 +/- 1 to 46 +/- 4 pg/ml (P less than 0.001) but did not alter mean serum 1,25-(OH)2D or urinary calcium or cyclic AMP in the nonobese subjects.(ABSTRACT TRUNCATED AT 250 WORDS)

The patency rate of internal mammary artery grafts is reported to be better than that of saphenous vein grafts in myocardial revascularization operations. To identify a possible biochemical explanation for this phenomenon, we studied the production of prostacyclin by the internal mammary artery and saphenous vein in 11 patients. Segments of internal mammary artery and saphenous vein from each patient were incubated in Krebs-Henseleit buffer at 37 degrees C. After 15 minutes, the basal production of 6-keto-prostaglandin F1 alpha (prostacyclin metabolite) by the internal mammary artery was 152 +/- 39 pg/mg wet weight (mean +/- standard error of the mean), whereas the saphenous vein produced only 68 +/- 17 pg/mg (p less than 0.001). After 30 minutes, the internal mammary artery produced 179 +/- 42 pg/mg, whereas the saphenous vein produced 75 +/- 18 pg/mg (p less than 0.001). After the basal incubation period, the vessels were incubated with arachidonic acid (prostaglandin substrate) for 15 minutes. The internal mammary artery produced 49.4 +/- 9.9 pg/mg, whereas the saphenous vein produced only 22.6 +/- 9.8 pg/mg (p less than 0.01). These observations suggest that the capacity of the internal mammary artery to produce prostacyclin in both a basal and a stimulated state is greater than that of the saphenous vein. Since prostacyclin is a potent vasodilator and inhibitor of platelet function, these results provide a possible biochemical explanation for the clinically observed better patency rate of internal mammary artery grafts.

1 The effects of pretreatment with the thromboxane synthetase inhibitor UK 37248 (dazoxiben) administered 30 min before intravenous endotoxin (S. enteriditis) in the rat was investigated 2 Plasma prostaglandins and thromboxanes were determined via radioimmunoassay. Endotoxaemia was associated with significant elevations above control values (less than 200 pg/ml) in plasma thromboxane B2 (TXB2), prostaglandin E (PGE) and 6-keto-prostaglandin F1 alpha (6-keto-PGF1 alpha). Within 30 min after endotoxin administration plasma immunoreactive (i) iTXB2 was 875 +/- 90 pg/ml (n = 9), iPGE was 1670 +/- 271 (n = 9) and i6-keto-PGF1 alpha was 1191 +/- 209 pg/ml (n = 10). By 4 h plasma iTXB2 was 1743 +/- 328 pg/ml (n = 5), iPGE was 2589 +/- 494 pg/ml (n = 9) and i6-keto-PGF1 alpha was 4251 +/- 984 pg/ml (n = 10). UK 37248 pretreatment resulted in a significant (P less than 0.001) decrease in plasma iTXB2 at 30 min and 4 h to 193 +/- 28 pg/ml (n = 5) and 421 +/- 57 pg/ml (n = 5), respectively. Unexpectedly UK 37248 also significantly decreased plasma i6-keto PGF1 alpha at 30 min and 4 h to 360 +/- 75 pg/ml (n = 10) (P less than 0.005) and 1920 +/- 513 pg/ml (n = 10) (P less than 0.05), respectively, iPGE plasma levels were not significantly changed in the UK 37248-pretreated rats 30 min (2210 +/- 370 pg/ml (n = 9) or 4 h 3529 +/- 1093 pg/ml (n = 13) after endotoxin compared to the vehicle-treated rats. 3 UK 37248 significantly (P less than 0.05) reduced the endotoxin mortality rate at 24 h from 69% (n = 13) to 30% (n = 13), UK 37248 also reduced splanchnic infarction from 90% (n = 20) to 6% (n = 16). 4 UK 37248 significantly improved the endotoxin-induced thrombocytopaenia, disseminated intravascular coagulation, hypoglycaemia and lysosomal labilization. 5 We conclude that UK 37248 provides significant beneficial effects in experimental endotoxic shock in the rat.

The objective of this study was to determine the relationships between the total oral clearance of propranolol and the partial clearances through its primary metabolic pathways, i.e. glucuronidation, side-chain oxidation and ring oxidation. Seven young, white males were given single 80 mg oral doses of the drug together with tritium-labelled propranolol. Plasma propranolol was measured by GC/MS and fourteen metabolites were measured in urine by h.p.l.c. with radiometric detection. The total oral clearance of propranolol in these subjects varied about three-fold, from 27.5 to 71.4 ml min-1 kg-1. The clearance through glucuronidation was very similar in all subjects, ranging from 6.8 to 9.9 ml min-1 kg-1. The clearance through side-chain oxidation varied 2.4-fold, from 10.9 to 25.8 ml min-1 kg-1. Increased clearance through this pathway correlated with increased total oral clearance (r = 0.84; P less than 0.02). The clearance through ring oxidation varied as much as 5.6-fold, from 7.5 to 41.8 ml min-1 kg-1. Increased clearance through this pathway correlated highly with increased total oral clearance (r = 0.94; P less than 0.002). These observations indicate that the intersubject variability in the oral clearance of propranolol in young, white males is due mainly to differences in the activity of the ring oxidation pathway, to some extent in the side-chain oxidation pathway, but not to differences in the glucuronidation pathway. The partial metabolic clearance approach adopted in this study may be useful in the investigation of factors influencing the oral bioavailability of propranolol.

We investigated the effects of the thromboxane synthetase inhibitor 7-(1-imidazolyl)heptanoic acid (7-IHA) and the fatty acid cyclooxygenase inhibitors indomethacin or ibuprofen in the treatment of fecal peritonitis in the rat. The effects of gentamicin alone and in combination with reduction of arachidonic acid metabolism by either treatment with indomethacin or essential fatty acid deficiency was also investigated. 7-IHA (60 mg/kg), administered i.p. 30 min before i.p. instillation of a fecal suspension, significantly reduced the plasma levels of immunoreactive (i) TxB2 from 1066 +/- 194 pg/ml (N = 14) to nondetectable (less than 200 pg/ml; N = 9) (P less than .01) at 1 hr and from 1695 +/- 218 (N = 16) to 508 +/- 56 pg/ml (N = 6) (P less than .01) at 4 hr after instillation of feces. In contrast, the levels of i6-keto-prostaglandin (PG)F1 alpha, the stable metabolite of prostacyclin, were significantly elevated by 7-IHA pretreatment from vehicle-treated septic control levels of 3777 +/- 414 (N = 16) to 5185 +/- 467 pg/ml (N = 9) (P less than .05) at 1 hr. Plasma i6-keto-PGF1 alpha at 4 hr in 7-IHA-treated rats (5503 +/- 665 pg/ml) (N = 6) was not different from vehicle-treated controls. Survival associated with fecal peritonitis was not altered by 7-IHA pretreatment. Indomethacin (10 mg/kg) or ibuprofen (5 mg/kg) administered i.p. 30 min before the fecal suspension significantly decreased both iTxB2 and i6-keto-PGF1 alpha, plasma levels when measured at 4 hr and prolonged survival time (P less than .05). Fibrinogen/fibrin degradation products were elevated (P less than .01) during fecal peritonitis and were reduced by indomethacin (P less than .01) or 7-IHA (P less than .05). Gentamicin significantly increased mean survival time from 8.6 +/- 0.2 (N = 50) to 23.8 +/- 2.6 hr (N = 16) (P less than .01). Gentamicin in combination with indomethacin or essential fatty acid deficiency further improved mean survival time and resulted in long-term survivals (greater than 48 hr) of 35 (N = 17) and 30% (N = 7), respectively (P less than .01 compared with gentamicin). Gentamicin pretreatment did not significantly alter plasma iTxB2 levels, but decreased i6-keto-PGF1 alpha from 9465 +/- 792 (N = 7) to 3096 +/- 1,174 pg/ml (N = 5; P less than .01) at 6 hr after induction of fecal peritonitis. These studies raise the possibility that inhibition of fatty acid cyclooxygenase may be a useful adjunct to antibiotic therapy in the treatment of septic shock.

Silymarin, an extract of crushed achenes of the milk thistle plant Silybum marianum is a multi-constituent mixture, 70-80% of which consists of a complex assortment containing the flavonolignans silybin A and B, isosilybin A and B, silydianin, and silychristin, and the flavonoid taxifolin. To date, numerous pharmacological actions of the silymarin extract have been documented in the biomedical literature, including hepatoprotective, anti-inflammatory, anti-tumor, and anti-fibrotic activities. The present study describes a novel liquid chromatographic-tandem mass spectrometric method for simultaneous analysis of silychristin, silydianin, silybin A and silybin B, isosilybin A and isosilybin B, and taxifolin in human plasma employing liquid-liquid extraction. This assay provides excellent resolution of the individual silymarin constituents via utilization of a 100 A 250 mm × 2 mm, 5 μm C(18) column with the mobile phase consisting of 51% methanol, 0.1% formic acid, and 10mM ammonium acetate. The lower limit of quantification was 2 ng/ml for each constituent. Calibration curves were linear over the range from 2 ng/ml to 100 ng/ml for all analytes (r(2)>0.99). The intra- and inter-day accuracies were 91-106.5% and 95.1-111.9%, respectively. The intra- and inter-day precision was within 10.5%. Additionally, recovery, stability, and matrix effects were fully validated as well. This method was successfully applied to human plasma samples from subjects treated with the milk thistle extract Legalon(®).

The renal kallikrein-kinin system is thought to be involved in vasoregulatory and epithelial ion-transporting processes. Renal kallikrein has not been studied in patients with diabetes mellitus, a disease in which abnormalities of renal hemodynamics and electrolyte handling occur. The urinary excretion of this kallikrein was measured in 20 type I diabetic patients and 10 normal subjects. On a 120-meq Na diet, daily kallikrein excretion, determined by both esterase activity and direct RIA, in 12 poorly controlled diabetic patients [hemoglobin A1c (HbA1c) = 14.2 +/- 0.5% (mean +/- SEM)] was significantly greater (P less than 0.05) than excretion in 8 diabetic patients in good to moderately good control (HbA1c = 9.4 +/- 0.5%) or in 10 normal subjects. In these groups, urinary esterase activities were 9.4 +/- 1.0, 6.1 +/- 1.4, and 6.7 +/- 0.5 esterase units/24 h, respectively. Corresponding excretion values of immunoreactive kallikrein were 171 +/- 14, 118 +/- 26, and 123 +/- 11 micrograms/24 h. Creatinine clearances were similar in the three groups. Urinary kallikrein was also measured in 8 diabetic and 8 normal subjects during 7 subsequent days of 10 meq Na intake. It increased less in diabetic patients than in normal subjects during Na depletion (P less than 0.02). The increase in urinary kallikrein in the diabetic patients was inversely related to their HbA1c levels (r = 0.88; P less than 0.01). The effect of glycemic control on urinary kallikrein excretion was determined in nine diabetic patients. Initial glycemic control was achieved using an artificial endocrine pancreas (Biostator) and was maintained by continuous sc insulin infusion with a portable pump. Before glycemic control, urinary kallikrein was 190 +/- 30 micrograms/24 h (by RIA). After 8-12 days of glycemic control, excretion fell to 144 +/- 23 micrograms/24 h (P less than 0.02). The abnormalities in kallikrein excretion in diabetic patients were not correlated with differences in water, electrolyte, protein, glucose, or aldosterone excretion in any of the studies. These results show that kallikrein excretion was increased in patients with poorly controlled insulin-dependent diabetes, and excretion rose less in diabetic subjects with low Na intake than in normal subjects. Strict glycemic control decreased urinary kallikrein excretion. These findings suggest that the renal kallikrein-kinin system is functioning abnormally in diabetes mellitus.

Human urinary kallikrein and an antiserum to it raised in the rabbit were used to detect and quantitate immunoreactive tissue kallikrein in human serum. Both 125I-labeled kallikrein and the unlabeled purified enzyme appear complexed to higher molecular weight entities in serum, but specific binding between radiolabeled enzyme and antiserum was unaffected by the presence of serum or plasma. Parallelism to standard displacement curves was always seen with radioimmunoassay of normal sera as well as with human mixed saliva or pancreatic extracts. Assay sensitivity is 160 pg/ml of serum, or 16 pg per tube. Purified plasma kallikrein or prekallikrein in concentrations up to 10 micrograms/ml showed no displacement. Acetone-kaolin activation of plasma produced the expected 30-fold increase in Tos-Arg-OMe esterase activity but no change in immunoreactive tissue kallikrein levels. Serum concentrations were 3.8 +/- 0.7 (mean +/- SE) ng/ml in 21 normal volunteers, and were similar in patients with Fletcher trait or Hageman factor deficiency. Significantly increased serum concentrations were seen with long-term low dietary sodium intake or acute forms of pancreatitis. Although the relation of this immunoreactive material to any active tissue kallikrein within the circulation remains to be determined, our studies provide a new parameter for the assessment of a system repeatedly suggested to have some role in regulation of vascular resistance.

Systemic lupus erythematosus (SLE) is an autoimmune disease that is characterized by the production of autoantibodies against nuclear antigens such as double-stranded DNA. Lupus predominantly affects women (ratio, 9:1). Moreover, premenopausal women with SLE are 50 times more likely to have a myocardial infarction. Although specific risk factors for advanced cardiovascular complications have not been identified in this patient population, endothelial dysfunction is highly prevalent. Recent studies show that the type I interferon signature gene expression coincides with impaired brachial artery flow-mediated dilation and diminished endothelial progenitor cell circulation, both markers of impaired endothelial function. Although many factors promote the development of vascular endothelial dysfunction, all pathways converge on the diminished activity of endothelial nitric oxide synthase (eNOS) and loss of nitric oxide (NO) bioavailability. Studies examining the effects of type I interferons on eNOS and NO in SLE are missing. This literature review examines the current literature regarding the role of type I interferons in cardiovascular disease and its known effects on regulators of eNOS and NO bioavailability that are important for proper endothelial cell function.

Platelets obtained from some diabetic patients show enhanced in vitro platelet aggregation. These studies were designed to determine if platelets obtained from diabetic subjects manifest increased metabolism of arachidonic acid to labile aggregating substances, such as thromboxane A2 (TXA2), and if they play a role in the enhanced platelet aggregation. Arachidonic acid stimulated TXA2 synthesis, as determined via radioimmunoassay of its stable metabolite TXB2, was significantly greater (p less than 0.01, n = 12) in platelet-rich plasma obtained from diabetic compared to matched controls. Arachidonic acid stimulated TXB2 synthesis in the diabetic platelet-rich plasma was positively correlated with the ambient fasting plasma glucose (r = 0.61, p less than 0.02, n = 15). Platelet aggregation induced by arachidonic acid (0.4-0.8 mM) was inhibited significantly less by 13-azaprostanoic acid (p less than 0.04, n = 14), an antagonist of the actions of prostaglandin H2 or TXA2 on platelets, compared to matched controls. We conclude that platelets obtained from some diabetic subjects manifest increased metabolism of arachidonic acid to labile aggregating substances which may contribute to the enhanced platelet aggregation.

The effects of thromboxane B(2) and the stable prostaglandin endoperoxide analogs (15Z)-hydroxy - 9alpha - 11alpha - (epoxymethano)prosta - 5Z,13E - dienoic acid (U44069) and (15Z)-hydroxy -11alpha,9alpha-(epoxymethano) prosta-5Z,13E-dienoic acid (U46619) were tested on water flow across the toad urinary bladder. In the presence of indomethacin or meclofenamic acid, inhibitors of prostaglandin and thromboxane A(2) synthesis, thromboxane B(2) stimulated water flow in a dose-dependent manner. U44069 (1 muM) stimulated water flow from 3.6+/-0.8 to 12.4+/-1.2 mg/min per 10 cm(2) hemibladder surface area, while U46619 (1 muM) stimulated water flow from 2.8+/-1.0 to 21.8+/-2.0 mg/min per 10 cm(2). The prostaglandin endoperoxide/thromboxane A(2) antagonist trans- 13-azaprostanoic acid, an inhibitor of vasopressin-stimulated water flow, inhibited thromboxane B(2)- and U46619-stimulated water flow in a dose-dependent manner. The inactive cis-13-azaprostanoic acid did not inhibit vasopressin-stimulated water flow in untreated hemibladders and had no effect on U46619-stimulated water flow in indomethacin or meclofenamic acid pretreated hemibladders. U46619 (1 muM) enhanced vasopressin-stimulated water flow in indomethacin pretreated hemibladders, producing a significant parallel shift (P < 0.001) in the dose-response relationship to submaximal concentrations of vasopressin (0.1-0.6 mU/ml), while not affecting water flow stimulated by supramaximal concentrations of vasopressin (10 mU/ml). trans-13-Azaprostanoic acid abolished the potentiating effects of U46619 on vasopressin-stimulated water flow. These results show that thromboxane A(2)-like compounds stimulate water flow in the toad urinary bladder.

OBJECTIVE

Lupus nephritis (LN) is an immune complex-mediated glomerulonephritis. Proliferative LN (PLN, ISN/RPS classes III and IV)) often leads to renal injury or failure despite traditional induction and maintenance therapy. Successful targeted therapeutic development requires insight into mediators of inflammation in PLN. Superoxide (SO) and its metabolites are mediators of the innate immune response through their ability to mediate reduction-oxidation signaling. Endothelial nitric oxide synthase (eNOS) modulates inflammatory responses in endothelial cells. We hypothesized that markers of SO production would be increased in active PLN and that SO production would be dependent on the activity of select enzymes in the renal cortex.

METHODS

Patients with systemic lupus erythematosus were enrolled at the time of renal biopsy for active LN of all classes. Serum collected at baseline was analyzed by HPLC with electrochemical detection for markers of SO production (durable modifications of serum protein Tyr ultimately requiring SO as a substrate). Renal cortex from MRL/MpJ-FAS(lpr) (MRL/lpr) mice with and without functional eNOS was analyzed during active disease for superoxide (SO) production with and without inhibitors of SO-producing enzymes.

RESULTS

Serum protein modifications indicative of total SO production were significantly higher in patients with PLN. These markers were increased in association with more active, inflammatory PLN. Mice lacking functional eNOS had 80% higher levels of renal cortical SO during active disease, and inhibitors of nitric oxide synthase and NADPH oxidase reduced these levels by 60% and 77%, respectively.

CONCLUSIONS

These studies demonstrate that SO production is unique to active PLN in a NOS and NADPH oxidase-dependent fashion. These findings suggest the emulating or augmenting eNOS activity or inhibiting NADPH oxidase SO production may be targets of therapy in patients with PLN. The markers of SO production used in this study could rationally be used to select SO-modulating therapies and serve as pharmacodynamic indicators for dose titration.

The effects of the organic calcium antagonists verapamil and methoxyverapamil (D-600) on vasopressin-stimulated water flow in response to an osmotic gradient were investigated. When toad bladders were exposed to either verapamil or D-600 for 60 min, vasopressin-stimulated water flow was inhibited. In the presence of verapamil (100 microM), water flow decreased from 38 +/- 3 to 26 +/- 3 mg/min/hemibladder (P less than .05, n = 5) and in the presence of D-600 (100 microM) from 48 +/- 3 to 32 +/- 4 mg/min/hemibladder (P less than .02, n = 5). When preincubated with hemibladders for 90 min they also inhibited basal 45Ca uptake into isolated toad bladder epithelial cells (from 1.58 +/- 0.20 to 1.02 +/- 0.14 nmol/mg of protein per 5 min, P less than .05, n = 6). Addition of verapamil to the bathing media during recovery from a vasopressin stimulus to block reuptake of calcium into the epithelial cells inhibited water flow in response to a subsequent incubation with vasopressin to a greater extent than the inhibition of water flow when verapamil was present only before exposure to vasopressin. Inhibition was also greater in the presence of lower extracellular calcium concentrations. The results are consistent with the notion that the calcium antagonists inhibit vasopressin-stimulated water flow by virtue of depleting a critical pool of intracellular calcium rather than preventing the simultaneous entry of calcium from extracellular sites during the exposure to vasopressin.